University of Groningen
Monitoring endothelial cells in microfluidic systems Grajewski, Maciej
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Publication date: 2018
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Grajewski, M. (2018). Monitoring endothelial cells in microfluidic systems. Rijksuniversiteit Groningen.
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Chapter 1
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Chapter 1
General introduction and the outline of the thesis
1.1 General introduction
The leading cause of premature death among citizens of developed countries is Cardiovascular Disease (CVD) [1], [2]. Endothelial dysfunction can be characteristic for different stages of progression for CVD [1], [3]. However, we currently lack the tools for monitoring those ample changes in the endothelium which lead to the development of a disease. It is therefore no surprise that the need for new ways to monitor the human endothelium is mentioned in World Health Organization reports on major causes of death in Western societies [1], [2]. This need has served as an important motivation for the work in this thesis, the focus of which is on the development of microsystems to study endothelial cell behavior.
The first challenge in this work was to understand how to prepare a suitable laboratory environment for cells that normally live in a human body. This environment needs to be adapted as much as possible to natural conditions to gain the best possible insight into the behavior of living cells taken out of a body. For this purpose, we employed microfluidic techniques to create microchannels with different dimensions to better represent the human vasculature in vitro. Additionally, we applied flow to recreate shear stress conditions, which are always present in an in vivo situation. In this way, we realized a microfluidic system for in vitro cell cultures which allows one to study the human endothelium under conditions which better correspond to those observed in microvasculature. Afterwards we present the development of an optical tool which can non-invasively monitor endothelial behavior in cell cultures. This project focused on a novel analytical approach and therefore required not just setting up the system but also understanding which phenomena we were observing and how to interpret them. It took some time before we understood that registered signals from the optical chip represent a phenomenon called cellular micromotion [4]. Micromotion is in essence the cytoskeletal rearrangement in response to external and internal stimuli, which results in changes in cell morphology. The potential of the optical chip in work with endothelial cells was verified in a number of experiments, which are described in this thesis.
The last part of this thesis revolved around 3D-printing. Throughout my PhD research, we benefited from using a 3D printer to facilitate experiments or improve the laboratory environment. Our experience has taught us about the possibilities and limitations of this technology. The application of 3D printing in daily laboratory work has been of great help to make my research easier.
All in all, my PhD research period has been an exciting journey both scientifically and personally for me. I successfully finished a number of research projects and acquired skills which allow me to say that I am a specialist in the field of microfluidics. The work performed within my PhD studies will positively contribute to different scientific endeavors in the next few years.
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1.2 References
[1] S. Mendis, P. Puska, and B. Norrving, Global Atlas on cardiovascular disease prevention and control., ISBN 978 92 4 156437 3; WHO, 2011.
[2] M. Ezzati, “Worldwide trends in blood pressure from 1975 to 2015 : a pooled analysis of 1479 population-based measurement studies with 19.1 million participants,” Lancet, vol. 389, pp. 37–55, 2017.
[3] W. C. Aird, “Endothelial cell heterogeneity.,” Cold Spring Harb. Perspect. Med., vol. 2, no. 1, pp. 1– 13, Jan. 2012.
[4] I. Giaever and C. R. Keese, “Micromotion of mammalian cells measured electrically.,” Proc. Natl. Acad. Sci. U. S. A., vol. 88, no. 17, pp. 7896–7900, 1991.
1.3 Outline of the thesis
In the first part of this thesis, we introduce the reader to the motivation for this work (Chapter 1) and methodology applied currently in cell culture research which served as an inspiration for the development of a new label-free approach in cell culture monitoring (Chapter 2). The second part of this work is dedicated to the presentation of an optimized microchannel design for microfluidic endothelial cell culture with detail protocols (Chapter 3). In Chapter 4, the continuation of the work with the optimized microchannel design is presented. In this Chapter endothelial cell cultures were submitted to different shear stresses, and occurring changes in protein distribution and cytoskeletal (re)arrangements in cells were observed with a confocal fluorescent microscope. The third part of this thesis describes the design, development and testing of a new label-free monitoring approach for endothelial cell culture, which works by propagating light through a few cells in a confluent cell culture (Chapter 5). In Chapter 6, xenobiotics which influence the cytoskeleton were added to endothelial cell cultures, and their effects were monitored with the developed optical chip. Chapter 7 is dedicated to the presentation of a versatile integrated cell culture platform, which was developed to improve performance of the optical chips developed in Chapter 5. Additionally, a microfluidic interface is added to the optical chip and tested. The fourth part of this thesis is dedicated to the application of 3D-printing technology in microfluidic research (Chapter 8). In this Chapter we present opportunities and limitation of 3D-printing technology based on our experience. The fifth part (Chapter 9) of this thesis discusses the impact of this work on the microfluidic field and strives to indicate future directions of development for microfluidic cell culture monitoring.
